The advent of "genomics" approaches has unveiled a diversity of phylogenetic mechanisms for generating immune receptor repertoires amongst metazoans. A lymphocyte antigen receptor system capable of generating enormous diversity via somatic rearrangement has recently been described in the agnathans (lamprey and hagfish), taxa representing the most primitive vertebrates but which lack the hallmark components necessary for adaptive immunity in higher vertebrates, i.e., the rearranging genes of the immunoglobulin superfamily (IgSF). These variable lymphocyte receptors (VLRs) are produced through an entirely novel molecular and genetic mechanism in which large banks of leucine rich repeat (LRR) cassettes are used to build the "diversity" region of the receptor molecules. These cassettes are imported into an incomplete germline gene from the other parts of the genome during lymphocyte development through an as-yet undefined mechanism. Based on its known sequence diversity, its inferred three-dimensional structure, and by implication to other known LRR-containing proteins such as the Toll-like receptors, the ectodomains of the VLRs are assumed to be capable of binding a wide range of pathogen specificities. Existence of this novel system suggests that there were at least two independent solutions to evolving an adaptive immune system in vertebrates and, in turn, raises many critical questions with regard to the evolutionary and genetic origins of the VLR system and our conception of adaptive immunity. Moreover, the discovery helps to validate the genomic approaches being embraced by the comparative immunology community in order to make inferences concerning the phylogeny of immunity. In this R21 application, we focus on two fundamental aspects of the VLR system, namely its role in early development and the genomic fine structure of its component modules. We will define the earliest stages in development at which the VLR system is operating, what cellular structures are involved, and whether it uses the same or different repertoire as previously defined. Moreover, we seek to determine whether the VLR system has an alternative role during early embryonic development. In terms of the genomic fine structure of its locus, we seek to genomically clone, characterize and sequence the entire VLR locus of the sea lamprey so as to better define how the complex is structured and how the rearrangements may be occurring. [unreadable] [unreadable] [unreadable]